Tissue samples were obtained from SBT specimens with fork lengths
of 114–130 cm during a normal commercial harvest from a tuna farm
located near Port Lincoln, South Australia. The fish were gilled and
gutted and the tail was removed. Samples of the tail muscle, liver, intestine
and kidney were frozen on dry ice. Upon return to the laboratory,
total lipid was extracted from the tissues using chloroform/methanol
(2:1 v/v) containing 0.05% (w/v) butylated hydroxyanisole using the
method described by Bligh and Dyer (1959) with some modifications
as described by Makrides et al. (1996). Subsamples of the total lipid
were separated into polar (mostly phospholipids) and neutral lipids
using thin layer chromatography (TLC) (Pahl et al., 2010). Briefly, the
subsamples were dried under a stream of nitrogen, reconstituted in
150 μL of chloroform/methanol (9:1 v/v) and then applied in streaks
to TLC plates. The plates were developed in petroleum spirit/diethyl
ether/acetic acid (180/30/2 by volume) and the separated lipid classes
were visualised by drying the plates, spraying them with fluorescein
5-isothiocyanate in methanol and then exposing them to UV light. The
bands of silica gel corresponding to the phospholipids were scraped
into glass vials containing 2 mL of 1% (v/v) H2SO4 in methanol and
FAME were produced by heating the vials to 70 °C for 3 h. Finally, the
FAME were extracted and analysed as previously described (Gregory
et al., 2011).